Chondrogenic differentiation during midfacial development in the mouse: in vivo and in vitro studies

Abstract

Because the mouse is now the main model for developmental research of all types, it is important to understand the basic developmental pattern of various organs. The first aim of the present study was to establish normal prenatal developmental standards of the cartilaginous nasal capsule during embryonic development of the mouse. For this purpose we have performed sagittal and coronal sections ranging from E12.5 to E18.5 in gestation age. The primordia of the nasal septal cartilage is recognizable around the 14th embryonic day as demonstrated by the metachromatic toluidine blue staining and by immunostaining of type II collagen. Northern blot analysis of the transcription factors Cart-1 and Sox-9 indicated maximum mRNA levels at E12.5 then a decreased expression during the following days of gestation. Type II collagen and aggrecan mRNA levels are constant during the embryonic period. In the second part of this study, we have established a primary culture system where chondrocytes were isolated from E.18 mouse embryo nasal septum. The purpose of this second part was to assess if chondrocytes could further differentiate in vitro until the hypertrophic phase and matrix mineralization. After the condensation phase, the cells synthesize an extracellular matrix including type II collagen and aggrecan. Progressively, typical cartilaginous nodules composed of clusters of round cells are visible, then increase in size and finally mineralize at day 12 of culture. Cart-1 and Sox-9 mRNA levels remain constant throughout the cultures, whereas type II collagen and aggrecan gradually decrease. Ultrastructural observations of the nodules show typical chondrocytes embedded in a dense network of fibers with matrix vesicles and mineralized foci. Other ultrathin sections revealed the presence of chondrons, typical of hyaline cartilage. Results from this study provide useful tools to further investigate morphogenesis and differentiation of the cartilaginous nasal capsule, and could in the future serve as a basic developmental standard.